| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
* Unité de Biochimie de la Nutrition, Faculté d’Ingénierie Biologique, Agronomique et Environnementale, Université Catholique de Louvain, B-1348 Louvain-la-Neuve, Belgium
Département Productions et Nutrition Animales, Centre Wallon de Recherches Agronomiques, B-5030 Gembloux, Belgium
1 Corresponding author: larondelle{at}bnut.ucl.ac.be
 |
ABSTRACT |
|---|
 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
Key Words: biohydrogenation • milk fat depression • oil-seed • vitamin E
|
INTRODUCTION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
Many experiments have been carried out to increase the CLA content of milk by feeding cows oil-rich diets. Unfortunately, the addition of unsaturated fat to cow feed has the disadvantage of increasing milk fat susceptibility to oxidation (Palmquist et al., 1993). Furthermore, it may result in progressive changes in the rumen environment, leading to decreased fiber digestion (Palmquist, 1984). This is associated with altered rumen biohydrogenation characterized by a shift in major biohydrogenation pathways: decreased formation of trans-11 C18:1 (vaccenic acid) and increased formation trans-10 C18:1 in the rumen (Griinari and Bauman, 1999). Because of the importance of vaccenic acid as the precursor of milk fat rumenic acid in the mammary gland, rumenic acid content eventually decreases in milk (Bauman et al., 2000).
Reduced milk fat is another important side effect of high-fat diets. This phenomenon, referred to as milk fat depression (MFD), is typically observed with low-fiber (LF) diets including high levels of concentrates (either readily digestible carbohydrates or unsaturated fat; Bauman and Griinari, 2003). Several theories have been proposed to explain MFD (Bauman and Griinari, 2003). However, currently, the most supported theory involves trans-10, cis-12 CLA as a direct inhibitor of milk fat synthesis in the mammary gland.
Charmley and Nicholson (1994) studied the effect of fat source on milk fat composition of cows receiving different levels of dietary vitamin E. They observed that dietary fat from micronized soybeans decreased milk fat concentration. However, a high concentration of vitamin E supplement in the diet (8,000 IU/d) eliminated the fat-depressing effect. Similarly, Focant et al. (1998) showed that a huge dietary vitamin E supplement (9,600 IU/d) prevented the drop in milk fat yield, as well as resistance to oxidation, which was induced by the incorporation of unsaturated fat-rich extruded linseeds and rapeseeds. Recently, Kay et al. (2005) observed that addition of 10,000 IU of 
-tocopherol/d to a TMR increased milk fat content by 6%.
The alleviating effect of vitamin E on MFD suggests that it could minimize the formation of trans-10 isomers in the rumen, because MFD is associated with high levels of those fatty acids in milk. Prevention of the biohydrogenation shift would then allow maintenance of both milk fat concentration and elevated level of rumenic acid in milk when plant oils or oilseeds are included in the diet. The present study aims to examine the effect of dietary supplementation with vitamin E on the production of trans-10 C18:1 and on MFD induced by feeding a high-UFA, LF diet.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
). This amount was calculated to cover animal needs according to INRA standards (INRA, 1988). The linseed diet was a 50:50 forage-to-concentrate ratio on a DM basis and contained extruded linseed (1.86 kg/d) and linseed oil (190 g/d). It was conceived to favor the trans-11 to trans-10 shift (low structural value, high level of UFA). The linseed + vitamin E diet was similar to the linseed diet except that it was supplemented with 12,000 IU of unprotected vitamin E/d added to the concentrates in the form of a powder containing 50% all-rac--tocopheryl acetate (Roche Co., Brussels, Belgium). The dietary changes were made overnight. Cows were offered feed individually twice daily at 0830 and 1830 h. Water was available at all times. Cows were milked twice daily at 0730 and 1730 h.
|
Analytical Procedures
Milk samples were refrigerated at 4°C. A portion of each sample was used for fat (Gerber, 1938) and CP (N x 6.37) analysis (Commission des Communautés Européennes, 1985), and the rest of each sample was processed into butter and anhydrous milk fat to allow long-term storage. To obtain anhydrous milk fat, butter was frozen overnight and then melted and centrifuged at 400 x g at 50°C for 15 min. The upper phase was passed through filters (filter type 595, Schleicher & Schuell, Dassel, Germany) filled with anhydrous Na2SO4 as a desiccant at 60°C. The extracted fat was stored at –20°C under N2 until analysis for fatty acids. Fatty acids were methylated in a solution of KOH in methanol (0.1 N) at 75°C for 60 min, then in a solution of HCl in methanol (1.2 N) at 75°C for 15 min, and finally extracted in hexane. Fatty acid methyl esters were quantified by gas chromatography (GC Trace ThermoQuest, Thermo-Finnigan, Milan, Italy) using a fused silica capillary column (100 m x 0.25 mm i.d. x 0.2-µm film thickness, CP-Sil 88, Chrompack, Middelburg, The Netherlands). Injection was on column. Hydrogen was used as the carrier gas. Pressure was held constant at 160 kPa. The initial oven temperature was 80°C, increased 25°C/min to 175°C (held for 25 min), then increased at 10°C/min to 205°C (held for 10 min), and finally decreased 20°C/min to 80°C. The temperature of the flame ionization detector was maintained at 255°C. Hydrogen flow rate to the detector was 25 mL/min; airflow was 350 mL/min. Each peak, except those of trans-10 C18:1 and trans-6 to trans-8 C18:1, was identified and quantified through comparison with pure methyl ester standards (Alltech Associates, Deerfield, IL, except CLA isomers from Nu-Chek Prep, Inc., Elysian, MN). Because no commercial standard is available for trans-10 C18:1, the concentration of the corresponding peak was calculated through comparison with the trans-11 C18:1 peak area (by multiplying the concentration of trans-11 C18:1 by the trans-10 peak area-to-trans-11 peak area ratio). The same method has been applied for trans-6 to trans-8 C18:1, which coelute. The validity of this method has been verified by applying it to the peak of trans-9 C18:1 and comparing this calculated concentration with the trans-9 C18:1 concentration determined through the use of the appropriate standard. For the determination of the different CLA isomers, 50 mL of raw milk was mixed with 10 mL of 25% NH3 and left for 15 min at room temperature. The fatty acids were then extracted by adding first 50 mL of ethanol, then 60 mL of petroleum ether, and lastly 60 mL of pentane, shaking well in between. The organic phases were then washed 2 times with 60 mL of distilled water and dried over anhydrous Na2SO4; finally, organic solvents were evaporated. The fatty acids were methylated with NaOCH3 methanol as described in Yurawecz et al. (1998). The CLA analysis was performed on a Ag+-HPLC (Gilson, Villiers le Bel, France) with 3 columns put in series (250 mm x 4.6 mm i.d. x 5-µm particle size, silver ion-impregnated, Chromspher lipids, Chrompack) with UV detection at 233 nm. The carrier liquid was acetonitrile and hexane (0.1:99.9, vol:vol), the flow was 1.06 cm3/min, and the oven temperature was 25.1°C.
Silage samples were lyophilized. All feed samples were ground in a mill (1-mm screen, Retsch, Dusseldorf, Germany) and analyzed for DM, OM, CP (N x 6.25), ether extract (Commissiondes Communautés Européennes, 1985), NDF and ADF (Goering and Van Soest, 1970), and -tocopherol (Brubacher et al., 1985). Lipids were extracted following the method of Folch modified by Christie (1982). Methylation and gas chromatography analyses were performed as described for milk samples.
Statistical Analyses
Based on the measurements on d 21 of each period (milk yield, milk fat, milk protein, fatty acid, and CLA profiles in milk), ANOVA was performed for a replicated Latin square design using the GLM procedure of SAS (SAS Inst., Inc., Cary, NC) with the following model:
 |
where Yijkl is the dependent variable, µ is the overall mean, cowi(l) is the cow effect inside each square (i = 1, 2, 3, 4, 5, or 6), dietj is the effect of treatments (j = 1 or 2), periodk is the effect of each period (k = 1 or 2), squarel is the effect of square l (l = 1, 2, or 3), and eijkl is the experimental error. Differences were declared significant at P < 0.05.
To account for pretrial differences between cows, ANOVA using the same model was also performed on the differences between the values on d 21 of each period and the values obtained from the pretrial measurement for each dependent variable. The results obtained with this approach were similar to those of the statistical analysis just described.
The correlation between milk fat and milk trans-10 C18:1 content was tested using all of the individual milk samples with the CORR procedure of SAS (SAS Inst., Inc.).
For each group, a one-tailed t-test was performed between the data of d 21 and 7 and the data of d 42 and 21 to highlight the increase or decrease of trans-11 C18:1 and trans-10 C18:1 during the 2 experimental periods.
|
RESULTS AND DISCUSSION |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
. They were very similar between the 2 diets, as the only difference was the vitamin E supplement. Intakes of ADF and NDF were intentionally low. Combined with a high intake of UFA (931 g/d on average), it was intended to favor the shift in biohydrogenation patterns. Vitamin E intake with the linseed + vitamin E diet was approximately 25 times higher than with the linseed diet.
|
, milk fat was low with the linseed diet, as it is classically observed when cows are fed high doses of UFA in a LF diet (Griinari et al., 1998). The ANOVA showed that the addition of vitamin E did not influence milk and protein yields. In contrast, milk fat yield and content were significantly increased with vitamin E supplementation. As described earlier in this paper, Charmley and Nicholson (1994), Focant et al. (1998), and Kay et al. (2005) observed a similar effect with dietary vitamin E.
|
). This is characteristic of cows fed high-concentrate, LF diets (Piperova et al., 2000, 2002; Peterson et al., 2003) and indicates that the linseed diet induced a shift in the ruminal biohydrogenation process. These results are consistent with those reported by Griinari et al. (1998). These authors examined the effect of fatty acid supplementation and the level of dietary fiber on MFD. Their LF diet containing high levels of UFA, which can be compared with the linseed diet in the present experiment, induced MFD and led to comparable levels of trans-11 C18:1 (2.52% of total fatty acids), trans-10 C18:1 (2.90% of total fatty acids), and cis-9, trans-11 C18:2 (1.10% of total fatty acids) in milk, as compared with those obtained with the linseed diet.
|
). The supplementation of the linseed diet with vitamin E significantly decreased the level of trans-10 C18:1 in milk, whereas it did not significantly increase the levels of vaccenic and rumenic acids. However, the graphs presented in Figure 1 show that the sequence of administration of vitamin E may influence its effect. When vitamin E was added simultaneously with linseed in the first period (Group 1), the level of trans-11 C18:1 was maintained, but trans-10 C18:1 remained low. This resulted in a high percentage of rumenic acid among the CLA isomers and a low percentage of trans-10, cis-12 C18:2 (data not shown). In the second period, when Group 1 stopped receiving vitamin E, trans-11 C18:1 decreased (P < 0.05) and trans-10 C18:1 increased (P < 0.05). Without vitamin E supplementation in the first period (Group 2), the shift occurred as expected: trans-10 C18:1 rose sharply (P < 0.01) and trans-11 C18:1 decreased (P < 0.05). However, when vitamin E was added at the beginning of the second period, it seemed to have only a limited effect. The levels of trans-11 C18:1 did not return to normal, even though a slight decrease was observed for trans-10 C18:1 (P = 0.11). Thus, vitamin E seems to have an effect on the levels of trans-11 C18:1 only when it is incorporated in the diet simultaneously with the fat. Once the shift has occurred, vitamin E is no longer able to counteract completely this process. In terms of daily yields, vitamin E had a significantly positive effect on the production of vaccenic acid and rumenic acid, as well as a significantly negative effect on the production of trans-10 C18:1 (Table 4). This can be explained by the vitamin E effect on milk fat content and yield. Finally, few changes were observed regarding the proportion of the CLA isomers in milk fat (Table 5). Surprisingly, vitamin E decreased the proportion of the trans-7, cis-9 isomer.
|
|
presents the correlation between milk fat content and milk trans-10 C18:1 level (y = 3.6945x–0.124; R2 = 0.4475; P < 0.001). The same type of curve has been observed by Bauman and Griinari (2003). Figure 3 illustrates the inverse correlation between trans-10 C18:1 in milk and milk fat content in the form of a temporal figure. This inverse correlation supports the link between trans-10 C18:1 and MFD. Bauman and Griinari (2001) proposed that fatty acid intermediates produced by altered rumen biohydrogenation would be responsible for diet-induced MFD. Studies using postruminal infusion or intravenous administration of trans-10, cis-12 C18:2 have clearly demonstrated that this CLA isomer is a potent inhibitor of milk fat synthesis in dairy cows. However, Peterson et al. (2003) suggested that other biohydrogenation intermediates should also contribute to the low milk fat syndrome. Griinari et al. (1998) suggested earlier that trans-10 C18:1 itself also induces MFD. Unfortunately, because of the lack of pure trans-10 C18:1, there have been no studies to firmly confirm its role in diet-induced MFD.
|
|
So far, adding buffering agents to the diet is the only method proposed to prevent trans-10 isomer formation in the rumen and consequent MFD (Piperova et al., 2002). Unfortunately, dietary buffers will also decrease overall production of trans-C18:1 fatty acids in the rumen (Kalscheur et al., 1997); therefore, less vaccenic acid may be available in the mammary gland to be desaturated into rumenic acid.
Factors resulting in the shift of biohydrogenation pathways are well known (LF diets supplied with high levels of UFA or readily digestive carbohydrates). However, the mechanism through which it occurs is less clear. Rumen bacteria are sensitive to the content of plant oil in the diet, because the UFA contained in these oils have an unspecified toxic effect on the rumen bacteria. Biohydrogenation could be the mechanism through which rumen bacteria cope with the presence of UFA in their environment (Harfoot and Hazlewood, 1988). However, when the load of UFA exceeds the biohydrogenation capacity, the growth as well as the function of rumen bacteria are impaired. Kim et al. (2002) showed that Megasphaera elsdenii, a bacterium producing trans-10 isomers, is more resistant to linoleic acid than Butyrivibrio fibrisolvens, a fiber-digesting bacterium that produces trans-11 isomers. Therefore, high-fat diets may be more detrimental to bacteria producing trans-11 isomers than those producing trans-10 isomers. Furthermore, high-concentrate, LF diets often result in increased lactic acid production, which eventually decreases ruminal pH (Griinari et al., 1998). The drop in ruminal pH may also contribute to altering ruminal microflora, leading to the shift in biohydrogenation pathways associated with LF diets. Fiber-digesting bacteria such as B. fibrisolvens would be sensitive to low pH, as opposed to M. elsdenii, which utilizes lactate (Kung and Hession, 1995).
Vitamin E may counteract the shift in the biohydrogenation pathways by supporting the growth and function of trans-11-producing bacteria. Hino et al. (1993) observed that addition of safflower oil to a culture of mixed rumen bacteria depressed the growth of rumen microbes. However, addition of -tocopherol and ß-carotene to the growth medium (each 5 mg/L) alleviated the growth inhibition by the UFA. Moreover, addition of -tocopherol and ß-carotene restored cellulose digestion, clearly by stimulating the growth of cellulolytic bacteria, which are supposed to be primarily responsible for the trans-11 biohydrogenation pathway (Martin and Jenkins, 2002).
The mechanism of vitamin E action is not known. In B. fibrisolvens, Hughes and Tove (1980a,b) identified compounds structurally close to -tocopherol—-to-copherolquinol and deoxy--tocopherolquinol—as electron donors in the biohydrogenation of rumenic acid to vaccenic acid. In that context, vitamin E may intervene in 3 different ways to help B. fibrisolvens to cope with an excess of UFA. First, vitamin E might act in place of the electron donors to provide the electrons for the reduction of the cis bond of the conjugated diene. Second, vitamin E might be metabolized to these donors by microorganisms in the rumen. However, Hughes and Tove (1980a) did not put forward such roles of -tocopherol in their experiment. Third, vitamin E might act as an electron donor to restore -tocopherolquinol and deoxy--tocopherolquinol. None of these hypotheses has been confirmed yet.
Another hypothesis would be that vitamin E inhibits the growth and function of bacteria that produce trans-10 C18:1. However, there is, at present, not much knowledge about this pathway; therefore, it is hard to say anything certain.
|
CONCLUSIONS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
|
ACKNOWLEDGEMENTS |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
Received for publication April 22, 2005. Accepted for publication October 14, 2005.
|
REFERENCES |
|---|
|
TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTS AND DISCUSSIONCONCLUSIONSACKNOWLEDGEMENTSREFERENCES |
|---|
-tocopherol on rumen bacteria in the utilization of long-chain fatty acids and cellulose. J. Dairy Sci. 76:600–605.-tocopherolquinol. J. Biol. Chem. 255:4447–4452.-tocopherolquinol as another endogenous electron donor for biohydrogenation. J. Biol. Chem. 255:11802–11806.This article has been cited by other articles:
|
|
 |
|
  M. Vazquez-Anon, J. Nocek, G. Bowman, T. Hampton, C. Atwell, P. Vazquez, and T. Jenkins Effects of Feeding a Dietary Antioxidant in Diets with Oxidized Fat on Lactation Performance and Antioxidant Status of the Cow J Dairy Sci, August 1, 2008; 91(8): 3165 - 3172. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. K. G. Kadegowda, L. S. Piperova, and R. A. Erdman Principal Component and Multivariate Analysis of Milk Long-Chain Fatty Acid Composition During Diet-Induced Milk Fat Depression J Dairy Sci, February 1, 2008; 91(2): 749 - 759. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
C. Hurtaud and J. L. Peyraud Effects of Feeding Camelina (Seeds or Meal) on Milk Fatty Acid Composition and Butter Spreadability J Dairy Sci, November 1, 2007; 90(11): 5134 - 5145. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. J. Moate, W. Chalupa, R. C. Boston, and I. J. Lean Milk Fatty Acids. I. Variation in the Concentration of Individual Fatty Acids in Bovine Milk J Dairy Sci, October 1, 2007; 90(10): 4730 - 4739. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Vazquez-Anon and T. Jenkins Effects of Feeding Oxidized Fat With or Without Dietary Antioxidants on Nutrient Digestibility, Microbial Nitrogen, and Fatty Acid Metabolism J Dairy Sci, September 1, 2007; 90(9): 4361 - 4367. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. E. Odongo, M. M. Or-Rashid, E. Kebreab, J. France, and B. W. McBride Effect of Supplementing Myristic Acid in Dairy Cow Rations on Ruminal Methanogenesis and Fatty Acid Profile in Milk J Dairy Sci, April 1, 2007; 90(4): 1851 - 1858. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
) in the milk of cows fed on a high-fat diet. Period 1 = d 1 to 21; Period 2 = d 22 to 42. Group 1 = Vitamin E added during Period 1; Group 2 = Vitamin E added during Period 2.
) and milk trans-10 C18:1 content (